Miniature pigs have been considered as a recommended large animal model for biomedical research. Mesenchymal stem cells offer promising potential for tissue regeneration. Recent studies have suggested that dental pulp stem cells (DPSCs) and periodontal ligament stem cells (PDLSCs) may provide more reliable strategies for the treatment of dental diseases using a cell-based tissue engineering approach. The aim of this study was to isolate and compare the characteristics of the DPSCs and PDLSCs of a miniature pig breed to the DPSCs and PDLSCs of a domestic farm pig breed. Stem cells of the DP and PDL were obtained from a male Yucatan miniature pig (nine months old) and a male domestic farm pig breed (six months old). The cell morphology, surface stem cell marker expression, proliferation, and osteogenic differentiation ability were evaluated. Under a light microscope, the DPSCs and PDLSCs of the miniature pig breed had morphologies similar to those of the domestic farm pig breed. The proliferation of PDLSCs in both animals showed no significant differences, except on day five, whereas the proliferation of DPSCs was significantly higher in the miniature pig breed. However, the osteogenic abilities of the DPSCs and PDLSCs from the miniature pig breed were significantly lower compared to the domestic farm pig breed. This observation emphasizes the need for the breed-specific optimization of an osteogenic differentiation culture protocol for Yucatan miniature pig DPSCs and PDLSCs before application to cell-based therapy for tissue engineering and regenerative medicine.